Studies on cytoplasmic components of sea urchin eggs stratified by centrifugation

Abstract

The participate components of the sea urchin egg cytoplasm stratified by centrifugation were studied in living and fixed material. The acetic iron, PAS and methyl green-pyronin stainings served as histochemical methods. The unfertilized eggs of Arbacia lixula, Echinocardium cordatum, Echinus esculentus, Paracentrotus lividus and Psammechinus miliaris were used as material. Sulphate esterified mucopolysaccharides are present in the cortex of all sea urchin eggs. The mode of distribution of the sulphate and α-glycol groupings varies according to the sea urchin species. This is in line with the species specific variations in the structure of cortical granules. The staining reactions of the cortical granules are summarized in Table I, p. 509. Sulphate esterified mucopolysaccharides are not only found in the cortex. The interior cytoplasm exhibits, after centrifugation, a layer of fine granules having the staining properties of acid mucopolysaccharides. There is a considerable variation with respect to the location and configuration of this layer in different sea urchin species. In the Arbacia egg the mucopolysaccharides are accumulated and precipitated as globules or, rather, flakes within the layer of fine granules, cf. Fig. 2 (layer c). In Echinocardium they form a narrow layer of granules of variable size enclosed in a hyaline matrix, cf. Figs. 7 and 9. In centrifuged Echinus eggs they appear as a discontinuous layer mixed with the centripetal region of the layer of yolk granules, cf. Fig. 18. In Psammechinus egg, however, they accumulate in the light layer ( a) as a uniform precipitate, cf. Fig. 18. Conversely, sections of the Paracentrotus eggs fails to reveal a layer of fine granules with the staining properties of mucopolysaccharides. The sulphate esterified polysaccharide complex does not seem to be bound to mitochondria. One more compound with the staining properties of acid mucopolysaccharides was found to be present in the yolk granules. In view of the chemically demonstrated presence of sialic acid in the yolk granules [24] it was suggested as possible that the spots stained with the acetic iron reagent contains sialic acid as a component. With respect to the relative density of yolk granules and ground cytoplasm of sea urchin eggs investigated two types of eggs are distinguished: (1) the eggs of Arbacia and Echinocardium with yolk granules which are heavy relative to the ground cytoplasm, and therefore accumulate centrifugally; (2) the eggs of Echinus, Paracentrotus and Psammechinus with yolk granules which are lighter relative to the ground cytoplasm. They therefore accumulate in a more centripetal or intermediary region. RNA and its derivatives are present in all layers of the stratified eggs. It was inferred, however, that in different layers of the centrifuged egg RNA may behave differently with respect to the combination with proteins and availability to staining reaction. A certain relation between the centrifugation and morphogenetic axes has been demonstrated in the egg of Echinocardium. The bearing of the histochemical results on the mechanism of formation of the fertilization membrane and on the determination of the dorso-ventral organization of the egg has been briefly discussed.